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Chemistry (Weinheim An Der Bergstrasse,... May 2024Biomolecules containing adenosine di- or triphosphate (ADP or ATP) are crucial for diverse biological processes. Synthesis of these biomolecules and development of their...
Biomolecules containing adenosine di- or triphosphate (ADP or ATP) are crucial for diverse biological processes. Synthesis of these biomolecules and development of their chemical probes are important to elucidate their functions. Enabling reproducible and high-yielding access to these ADP- and ATP-containing molecules via conventional P(III)-P(V) and P(V)-P(V) coupling reactions is challenging owing to water content in highly polar phosphate-containing substrates. Herein, we report an efficient and reliable method for protecting-group-free P(V)-P(V) coupling reaction through in situ activation of phosphates using hydrolysis-stable 2-[N-(2-methylimidazoyl)]-1,3-dimethylimidazolinium chloride (2-MeImIm-Cl), providing the corresponding electrophilic P(V) intermediates for subsequent nucleophilic attack using their coupling partners. This P(V)-P(V) coupling reaction proceeded even in a wet reaction medium and showed a broad substrate scope, accommodating protecting-group-free synthesis of ADP-ribose and nicotinamide adenine diphosphate analogs, ATP-containing biomolecules, and ADP-ribosyl peptides.
PubMed: 38763895
DOI: 10.1002/chem.202401302 -
Bioorganic Chemistry Jul 2024Adenylate kinase (AK) plays a crucial role in the metabolic monitoring of cellular adenine nucleotide homeostasis by catalyzing the reversible transfer of a phosphate...
Adenylate kinase (AK) plays a crucial role in the metabolic monitoring of cellular adenine nucleotide homeostasis by catalyzing the reversible transfer of a phosphate group between ATP and AMP, yielding two ADP molecules. By regulating the nucleotide levels and energy metabolism, the enzyme is considered a disease modifier and potential therapeutic target for various human diseases, including malignancies and inflammatory and neurodegenerative disorders. However, lacking approved drugs targeting AK hinders broad studies on this enzyme's pathological importance and therapeutic potential. In this work, we determined the effect of a series of dinucleoside polyphosphate derivatives, commercially available (11 compounds) and newly synthesized (8 compounds), on the catalytic activity of human adenylate kinase isoenzyme 1 (hAK1). The tested compounds belonged to the following groups: (1) diadenosine polyphosphates with different phosphate chain lengths, (2) base-modified derivatives, and (3) phosphate-modified derivatives. We found that all the investigated compounds inhibited the catalytic activity of hAK1, yet with different efficiencies. Three dinucleoside polyphosphates showed IC values below 1 µM, and the most significant inhibitory effect was observed for P-(5'-adenosyl) P-(5'-adenosyl) pentaphosphate (ApA). To understand the observed differences in the inhibition efficiency of the tested dinucleoside polyphosphates, the molecular docking of these compounds to hAK1 was performed. Finally, we conducted a quantitative structure-activity relationship (QSAR) analysis to establish a computational prediction model for hAK1 modulators. Two PLS-regression-based models were built using kinetic data obtained from the AK1 activity analysis performed in both directions of the enzymatic reaction. Model 1 (AMP and ATP synthesis) had a good prediction power (R = 0.931, Q = 0.854, and MAE = 0.286), while Model 2 (ADP synthesis) exhibited a moderate quality (R = 0.913, Q = 0.848, and MAE = 0.370). These studies can help better understand the interactions between dinucleoside polyphosphates and adenylate kinase to attain more effective and selective inhibitors in the future.
Topics: Humans; Quantitative Structure-Activity Relationship; Dinucleoside Phosphates; Kinetics; Molecular Structure; Adenylate Kinase; Dose-Response Relationship, Drug; Protein Kinase Inhibitors; Enzyme Inhibitors
PubMed: 38744169
DOI: 10.1016/j.bioorg.2024.107432 -
Nature Communications May 2024c-di-AMP is an essential and widespread nucleotide second messenger in bacterial signaling. For most c-di-AMP synthesizing organisms, c-di-AMP homeostasis and the...
c-di-AMP is an essential and widespread nucleotide second messenger in bacterial signaling. For most c-di-AMP synthesizing organisms, c-di-AMP homeostasis and the molecular mechanisms pertaining to its signal transduction are of great concern. Here we show that c-di-AMP binds the N-acetylglucosamine (GlcNAc)-sensing regulator DasR, indicating a direct link between c-di-AMP and GlcNAc signaling. Beyond its foundational role in cell-surface structure, GlcNAc is attractive as a major nutrient and messenger molecule regulating multiple cellular processes from bacteria to humans. We show that increased c-di-AMP levels allosterically activate DasR as a master repressor of GlcNAc utilization, causing the shutdown of the DasR-mediated GlcNAc signaling cascade and leading to a consistent enhancement in the developmental transition and antibiotic production in Saccharopolyspora erythraea. The expression of disA, encoding diadenylate cyclase, is directly repressed by the regulator DasR in response to GlcNAc signaling, thus forming a self-sustaining transcriptional feedback loop for c-di-AMP synthesis. These findings shed light on the allosteric regulation by c-di-AMP, which appears to play a prominent role in global signal integration and c-di-AMP homeostasis in bacteria and is likely widespread in streptomycetes that produce c-di-AMP.
Topics: Acetylglucosamine; Allosteric Regulation; Signal Transduction; Bacterial Proteins; Gene Expression Regulation, Bacterial; Dinucleoside Phosphates; Saccharopolyspora
PubMed: 38714645
DOI: 10.1038/s41467-024-48063-0 -
Acta Crystallographica. Section D,... May 2024CdaA is the most widespread diadenylate cyclase in many bacterial species, including several multidrug-resistant human pathogens. The enzymatic product of CdaA, cyclic...
CdaA is the most widespread diadenylate cyclase in many bacterial species, including several multidrug-resistant human pathogens. The enzymatic product of CdaA, cyclic di-AMP, is a secondary messenger that is essential for the viability of many bacteria. Its absence in humans makes CdaA a very promising and attractive target for the development of new antibiotics. Here, the structural results are presented of a crystallographic fragment screen against CdaA from Listeria monocytogenes, a saprophytic Gram-positive bacterium and an opportunistic food-borne pathogen that can cause listeriosis in humans and animals. Two of the eight fragment molecules reported here were localized in the highly conserved ATP-binding site. These fragments could serve as potential starting points for the development of antibiotics against several CdaA-dependent bacterial species.
Topics: Listeria monocytogenes; Crystallography, X-Ray; Binding Sites; Bacterial Proteins; Models, Molecular; Dinucleoside Phosphates; Anti-Bacterial Agents; Humans; Phosphorus-Oxygen Lyases; Protein Conformation
PubMed: 38682668
DOI: 10.1107/S205979832400336X -
Proceedings of the National Academy of... Apr 2024In bacteria, intracellular K is involved in the regulation of membrane potential, cytosolic pH, and cell turgor as well as in spore germination, environmental...
In bacteria, intracellular K is involved in the regulation of membrane potential, cytosolic pH, and cell turgor as well as in spore germination, environmental adaptation, cell-to-cell communication in biofilms, antibiotic sensitivity, and infectivity. The second messenger cyclic-di-AMP (c-di-AMP) has a central role in modulating the intracellular K concentration in many bacterial species, controlling transcription and function of K channels and transporters. However, our understanding of how this regulatory network responds to c-di-AMP remains poor. We used the RCK (Regulator of Conductance of K) proteins that control the activity of Ktr channels in as a model system to analyze the regulatory function of c-di-AMP with a combination of in vivo and in vitro functional and structural characterization. We determined that the two RCK proteins (KtrA and KtrC) are neither physiologically redundant or functionally equivalent. KtrC is the physiologically dominant RCK protein in the regulation of Ktr channel activity. In explaining this hierarchical organization, we found that, unlike KtrA, KtrC is very sensitive to c-di-AMP inactivation and lack of c-di-AMP regulation results in RCK protein toxicity, most likely due to unregulated K flux. We also found that KtrC can assemble with KtrA, conferring c-di-AMP regulation to the functional KtrA/KtrC heteromers and potentially compensating KtrA toxicity. Altogether, we propose that the central role of c-di-AMP in the control of the K machinery, by modulating protein levels through gene transcription and by regulating protein activity, has determined the evolutionary selection of KtrC as the dominant RCK protein, shaping the hierarchical organization of regulatory components of the K machinery.
Topics: Bacterial Proteins; Bacillus subtilis; Potassium; Gene Expression Regulation, Bacterial; Dinucleoside Phosphates; Potassium Channels
PubMed: 38652747
DOI: 10.1073/pnas.2318666121 -
Journal of the American Chemical Society Apr 2024Nonenzymatic template-directed RNA copying requires catalysis by divalent metal ions. The primer extension reaction involves the attack of the primer 3'-hydroxyl on the...
Nonenzymatic template-directed RNA copying requires catalysis by divalent metal ions. The primer extension reaction involves the attack of the primer 3'-hydroxyl on the adjacent phosphate of a 5'-5'-imidazolium-bridged dinucleotide substrate. However, the nature of the interaction of the catalytic metal ion with the reaction center remains unclear. To explore the coordination of the catalytic metal ion with the imidazolium-bridged dinucleotide substrate, we examined catalysis by oxophilic and thiophilic metal ions with both diastereomers of phosphorothioate-modified substrates. We show that Mg and Cd exhibit opposite preferences for the two phosphorothioate substrate diastereomers, indicating a stereospecific interaction of the divalent cation with one of the nonbridging phosphorus substituents. High-resolution X-ray crystal structures of the products of primer extension with phosphorothioate substrates reveal the absolute stereochemistry of this interaction and indicate that catalysis by Mg involves inner-sphere coordination with the nonbridging phosphate oxygen in the pro- position, while thiophilic cadmium ions interact with sulfur in the same position, as in one of the two phosphorothioate substrates. These results collectively suggest that during nonenzymatic RNA primer extension with a 5'-5'-imidazolium-bridged dinucleotide substrate the interaction of the catalytic Mg ion with the pro- oxygen of the reactive phosphate plays a crucial role in the metal-catalyzed S2(P) reaction.
Topics: RNA; Metals; Dinucleoside Phosphates; Phosphates; Catalysis; Oxygen; Ions; RNA, Catalytic
PubMed: 38579124
DOI: 10.1021/jacs.4c00323 -
Organic & Biomolecular Chemistry Apr 2024Four dinucleotide analogs of thymidylyl(3'-5')thymidine (TpT) have been designed and synthesized with a view to increase the selectivity, with respect to CPD, of...
Four dinucleotide analogs of thymidylyl(3'-5')thymidine (TpT) have been designed and synthesized with a view to increase the selectivity, with respect to CPD, of efficient UV-induced (6-4) photoproduct formation. The deoxyribose residues of these analogs have been modified to increase north and south conformer populations at 5'- and 3'-ends, respectively. Dinucleotides whose 5'-end north population exceeds 60% and whose 3'-end population is almost completely south display a three-fold selective enhancement in (6-4) adduct production when exposed to UV radiation, compared to TpT. These experimental results undoubtedly provide robust foundations for studying the singular ground-state proreactive species involved in the (6-4) photoproduct formation mechanism.
Topics: Sugars; Photochemistry; Carbohydrates; Dinucleoside Phosphates; Ultraviolet Rays
PubMed: 38530278
DOI: 10.1039/d4ob00279b -
Biomaterials Science Apr 2024Colorectal cancer (CRC) ranks among the most prevalent cancers globally, demanding innovative therapeutic strategies. Immunotherapy, a promising avenue, employs cancer...
Colorectal cancer (CRC) ranks among the most prevalent cancers globally, demanding innovative therapeutic strategies. Immunotherapy, a promising avenue, employs cancer vaccines to activate the immune system against tumors. However, conventional approaches fall short of eliciting robust responses within the gastrointestinal (GI) tract, where CRC originates. Harnessing the potential of all-trans retinoic acid (ATRA) and cytosine-phosphorothioate-guanine (CpG), we developed layered nanoparticles using a layer-by-layer assembly method to co-deliver these agents. ATRA, crucial for gut immunity, was efficiently encapsulated alongside CpG within these nanoparticles. Administering these ATRA@CpG-NPs, combined with ovalbumin peptide (OVA), effectively inhibited orthotopic CRC growth in mice. Our approach leveraged the inherent benefits of ATRA and CpG, demonstrating superior efficacy in activating dendritic cells, imprinting T cells with gut-homing receptors, and inhibiting tumor growth. This mucosal adjuvant presents a promising strategy for CRC immunotherapy, showcasing the potential for targeting gut-associated immune responses in combating colorectal malignancies.
Topics: Tretinoin; Animals; Colorectal Neoplasms; Nanoparticles; Mice; Humans; Adjuvants, Immunologic; Mice, Inbred C57BL; Female; Immunotherapy; Ovalbumin; Cell Line, Tumor; Mice, Inbred BALB C; Oligodeoxyribonucleotides; Dendritic Cells; Layer-by-Layer Nanoparticles; Dinucleoside Phosphates
PubMed: 38498328
DOI: 10.1039/d4bm00026a -
BMJ Case Reports Feb 2024A man in his 50s presented in an emergency with breathlessness and chest discomfort. On evaluation, he was diagnosed with coronary artery disease, with more than 80%...
A man in his 50s presented in an emergency with breathlessness and chest discomfort. On evaluation, he was diagnosed with coronary artery disease, with more than 80% narrowing of the right coronary and left circumflex arteries. The patient underwent percutaneous coronary intervention and was started on dual antiplatelet (DAPT) therapy. After starting DAPT, the patient developed gross haematuria with a drop in haematocrit. Further evaluation revealed a left renal mass with urinary bladder clots. Because of the risk of stent thrombosis on stopping DAPT, radical nephrectomy was deferred, and the patient underwent left renal artery angioembolisation and bladder clot evacuation. On the follow-up, the patient was stable with a gradual decrease in renal mass size, and after a year, the patient underwent definitive surgery. The patient is doing well in 4 years of follow-up with no metastasis.
Topics: Humans; Male; Carcinoma, Renal Cell; Coronary Artery Disease; Dinucleoside Phosphates; Drug Therapy, Combination; Drug-Eluting Stents; Hemorrhage; Kidney Neoplasms; Myocardial Infarction; Platelet Aggregation Inhibitors; Thrombosis; Middle Aged
PubMed: 38423578
DOI: 10.1136/bcr-2023-257750 -
International Journal of Molecular... Dec 2023To improve the chemical regulation on the activity of cyclic dinucleotides (CDNs), we here designed a reduction-responsive dithioethanol (DTE)-based dCDN prodrug...
To improve the chemical regulation on the activity of cyclic dinucleotides (CDNs), we here designed a reduction-responsive dithioethanol (DTE)-based dCDN prodrug (DTE-dCDN). Prodrug improved the cell permeability with the intracellular levels peaking in 2 h in THP-1 cells. Under the reductive substance such as GSH or DTT, prodrug could be quickly decomposed in 30 min to release the parent dCDN. In THP1-Lucia cells, prodrug also retained a high bioactivity with the EC of 0.96 μM, which was 51-, 43-, and 3-fold more than the 2',3'-cGAMP (EC = 48.6 μM), the parent compound 3',3'-c-di-dAMP (EC = 41.3 μM), and ADU-S100 (EC = 2.9 μM). The high bioactivity of prodrug was validated to be highly correlated with the activation of the STING signaling pathway. Furthermore, prodrug could also improve the transcriptional expression levels of , , and in THP-1 cells. These results will be helpful to the development of chemically controllable CDN prodrugs with a high cellular permeability and potency.
Topics: Deoxyribose; Alarmins; Dinucleoside Phosphates; Permeability; Prodrugs
PubMed: 38203256
DOI: 10.3390/ijms25010086